Dry phase reagent strips incorporating enzyme-based compositions are used extensively in clinical laboratories, physician's offices, hospitals, and homes to test samples of biological fluids for glucose concentration. In fact, reagent strips have become an everyday necessity for many of the nation's several million diabetics. Since diabetes can cause dangerous anomalies in blood chemistry, it can contribute to vision loss and kidney failure. To minimize the risk of these consequences, most diabetics must test themselves periodically, then adjust their glucose concentration accordingly, for instance, through diet control and/or with insulin injections. Some patients must test their blood glucose concentration as often as four times daily or more.
It is especially important for diabetic individuals who must control their diet in order to regulate sugar intake and/or administer insulin injections, and who must be guided in this regard by frequent tests of blood glucose concentration, to have rapid, inexpensive, and accurate reagent strips for glucose determination.
Reagent strips are known that contain an oxidizable dye or indicator that turns a different shade of color, depending on the concentration of glucose in a biological fluid that has been applied to the reagent strip. Reagent strips are known which include an enzyme, such as glucose oxidase, which is capable of oxidizing glucose to gluconic acid lactone and hydrogen peroxide. Known reagent strips also contain an oxidizable dye and a substance having peroxidative activity which is capable of selectively catalyzing oxidation of the oxidizable dye in the presence of hydrogen peroxide. (See, for example, U.S. Pat. No. 5,306,623.)
Whether the test is conducted in the home, physician's office, clinic or a hospital, accuracy and reproducibility of the glucose determination are extremely important. In the case of a color-indicating reagent strip, it is desirable that the color change be pronounced and be insensitive to variations in compounds contained in the biological fluid other than glucose. In the case of a visually-read reagent strip, it is especially important that diabetics, who may be suffering from impaired vision, have a test reagent that exhibits a significant color change dependent upon glucose concentration. Color change, as exhibited by a change in absorbance and/or reflectance at a given wavelength, is also important for the accuracy of meter-read strips.
The performance of reagent strips may be affected by the presence of various interfering factors, and the need to reduce or eliminate the effect of interferents in clinical chemistry has been recognized by others. For example, Arter et al., EPO Application No. 93111290.8, published on Jan. 19, 1994, discloses an analytical element for the determination of an analyte in an aqueous fluid. The element includes a reagent capable of binding free sulfhydryl groups present in the fluid. Arter et al. are primarily interested in detecting analytes such as acetaminophen, salicylate, creatinine, cholesterol, HDL cholesterol, triglycerides, glucose and uric acid. They use enzymes that produce hydrogen peroxide, which is assayed by way of peroxidase-coupled redox chemistry. The interfering sulfhydryl groups (such as N-acetyl cysteine) may be present in the fluid and can trigger oxidation/reduction, even in the absence of the analyte. Arter et al. address the problem of the interfering free sulfhydryl groups by adding agents that react with and bind sulfhydryl groups. Suitable agents include maleimide, N-ethylmaleimide, iodoacetamide, silver nitrate, and gold chloride.
Ismail et al., U.S. Pat. No. 5,185,247, discloses an enzyme-based test strip that is stabilized by the addition of various compositions. He includes imidazole among the agents that impart heat stability, but it is only effective if the strip is also impregnated with an "ascorbate interference composition," which includes mercuric oxide and sarcosine. Incorporating heavy-metal oxides in the reagent is generally undesirable, because they are toxic.
It is known that interfering factors can affect the color change of reagent strips that measure blood glucose. For example, reduced color changes in glucose determinations seem to correlate generally with the concentration of red blood cells in the biological fluid sample (the hematocrit). Of course, any inaccuracy is magnified when the actual value of glucose concentration to be determined is low.
There is a need for a reagent strip that provides a pronounced change in color along a glucose-concentration continuum when exposed to biological fluids containing glucose. Further, there is a need for reagent test strips that are capable of providing reproducible results for glucose determination, regardless of fluctuations in the concentrations of other components found in the biological fluids. Ideally, a reagent strip result should remain invariant in its analysis of glucose concentration, even though exposed to whole blood samples (1) having hematocrit levels that vary throughout the range of 25% to 60% hematocrit and (2) containing other components commonly found in blood. Finally, it is desirable to have a reagent strip that does not incorporate heavy-metal oxide.